Lactosaminoglycan assembly, cell surface expression, and release by mouse uterine epithelial cells.
The kinetics of assembly, cell surface expression, secretion, and degradation of the major lactosaminoglycan (LAG)-bearing glycoproteins in mouse uterine epithelial cells have been studied. LAGs have been shown previously to be synthesized preferentially by these cells in the uterus and are expressed at the cell surface, where they participate in cell adhesion processes (Dutt, A., Tang., J.-P., and Carson, D. D. (1987) Dev. Biol. 119, 27-37). We utilized selection on pokeweed mitogen-Sepharose, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and subsequent electroelution to isolate the major LAG-bearing glycoproteins. The intact LAG-bearing glycoproteins exhibited very high apparent Mr (greater than 500,000) both by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and molecular exclusion chromatography under dissociative conditions. The subset of LAGs at the cell surface exhibited a half-life of approximately 11 h, whereas total cell-associated LAGs had a half-life of 6 +/- 1 h. Pulse-chase experiments indicated that the transit time of LAG core proteins from the rough endoplasmic reticulum to the site of LAG addition in the Golgi was 30-45 min. LAG glycoprotein transit from the Golgi to the cell surface required at least an additional 30-45 min. The major metabolic fate of the cell-associated LAGs was secretion to the medium with no evidence of lysosomal degradation. Some (30%) of the LAGs appeared to be released to the medium via the action of cell surface proteases. Epithelial cell surfaces bound fluoresceinated pokeweed mitogen, indicating the constitutive presence of LAG-bearing molecules at the cell surface; pokeweed mitogen binding to the cell surface was completely blocked by 10 mM chitotriose. These observations provide the first comprehensive description of the intracellular transport and metabolism of this interesting class of glycoproteins of the uterine epithelial cell surface.
Dutt
A
A
Department of Biochemistry and Molecular Biology, University of Texas, M.D. Anderson Cancer Center, Houston 77030.
Carson
D D
DD
D000606:Amino Sugars / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D005260:Female; D005944:Glucosamine / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D006207:Half-Life; D007700:Kinetics; D051379:Mice; D008970:Molecular Weight; D011043:Pokeweed Mitogens / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism*; D014599:Uterus / Q000378:metabolism* / Q000648:ultrastructure
D000606:Amino Sugars; D006023:Glycoproteins; D011043:Pokeweed Mitogens; D011134:Polysaccharides; C039448:lactosaminoglycan; D005944:Glucosamine